There are two distinct pathways of allorecognition by T cells. In the so-called "direct" pathway T cells recognize intact allo-MHC molecules on the surface of donor cells, including donor graft vascular endothelial cells. In the so-called "indirect" pathway T cells recognize processed alloantigen presented as peptides by self-APCs. Studies carried out in Dr. Briscoe?s laboratory have provided a foundation for the identification of a novel regulatory function of the endothelium in the indirect pathway of allorecognition. Briefly, when recipient monocytes transmigrate across allogeneic endothelium, they receive signals that result in alterations in cell surface phenotype, the expression of costimulatory molecules and cytokines; and differentiation into functional myeloid-derived dendritic cells. These extremely efficient APCs emerge from the graft into lymph nodes where they may function in the indirect pathway of allorecognition. In this research, proposal we will: 1) test the hypothesis that interactions between recipient monocytes (self-APCs) and allogeneic endothelial cells promote indirect T cell responses to alloantigen, 2) test the hypothesis that the critical function of the endothelium in indirect allorecognition is the recruitment and delivery of APCs into the graft; and that it is the degree of intragraft apoptosis and necrosis that regulates persistence of the indirect pathway; and 3) test the hypothesis that self-restricted presentation of allopeptide by endothelial cells augments the recruitment of indirectly primed T cells into allografts with chronic rejection. These studies will utilize T cells, APCs and endothelial cells from patients enrolled in Project 3 to enable us to address whether monocyte-endothelial cell interactions and resultant indirect responses are of functional significance for the development of chronic rejection. In addition, we will collaborate with Project 1 to assess if this mechanism(s) is functional in the animal model of chronic rejection; and we will utilize the scientific core and the administrative and data management core for our analyses. These combined interactions provide for a cohesive research strategy, in which these in vitro mechanistic studies can be addressed in the context of chronic rejection in vivo. Such knowledge has distinct clinical implications and suggests novel strategies to inhibit the indirect response and thus chronic rejection.